Project Summary: A major cause of diabetes is impairment of glucose uptake by skeletal muscle that causes pumping of extra glucose into blood leading to hyperglycemia. Clinical studies revealed that diabetes causes cardiomyopathy and the chances of heart failure increases if the patient has diabetes. One of the mechanisms of cardiac dysfunction associated with diabetes is oxidative stress that activates latent matrix metalloproteinase-9 (MMP9), which in turn induces fibrosis and contractile dysfunction. However, the specific mechanisms for how oxidative stress activates MMP9, which leads to contractile dysfunction, have not been investigated. Our preliminary studies on HL1 cardiomyocytes suggest that inhibition of miR-133 induces MMP9 and over expression of miR-133 inhibits MMP9. The luciferase reporter assay revealed that miR-133 targets MMP9. Interestingly, glucose mediated induction of MMP9 is abrogated by miR-133. In the heart of diabetic Akita mice, myosin enhancer factor 2c (Mef2c- an inducer of miR-133) is alleviated, miR-133 is down regulated and MMP9 is robust. These results lead us to hypothesize that oxidative stress inhibits Mef2c causing attenuation of miR-133 that induces MMP9 leading to contractile dysfunction in diabetes. To address the hypothesis, we formulated three specific aims: Aim#1: To determine whether the miR-133 directly or indirectly inhibits the activation of MMP9. Hypothesis: MiR-133 directly inhibits MMP9 by targeting its 3/ UTR. It also indirectly inhibits MMP9 by inducing miR-466 and abrogating miR-705. Aim# 2: To determine whether the oxidative stress inhibits Mef2c causing attenuation of miR-133 in diabetes. Hypothesis: The oxidative stress inhibits Mef2c that causes attenuation of miR-133 in diabetes. Aim # 3: To determine whether the over-expression of miR-133 or Mef2c will inhibit MMP9 that in turn improve glucose uptake in skeletal muscle and ameliorates contractile dysfunction in diabetes. Hypothesis: The over expression of miR-133 and Mef2c inhibits MMP9 that enhances glucose uptake by skeletal muscle and mitigates contractile dysfunction of cardiomyocytes in diabetes. Our proposal unravels a new mechanism of regulation of MMP9 by miR-133. It also provides a new concept that miRNA inhibits a gene not only by directly targeting it rather it also induces / inhibits other miRNAs that indirectly influences the target gene. At translational level, the proposal will provide concrete evidence that over expression of miR-133 or ablation of MMP9 can ameliorate diabetic cardiomyopathy.